Follicular helper T (TFH) cells are critical for development of germinal center (GC) B cells responses following immunization with TD antigens and in systemic autoimmune responses;however, the development and function of these cells have been less clearly defined in both normal individuals and in patients with systemic lupus erythematosus than other CD4 T cell effector subsets. These issues have been a focus of study for the past several years, with characterization of CD4 T-B helper cells in murine models of lupus and in conventional immune responses, with the idea that information gleaned from the latter studies could be applied to the understanding of autoimmunity. The proposed work builds on these observations, and the more recent finding that TFH cells require the expression of the transcriptional repressor Bcl6 (B-cell CLL/lymphoma 6) for their development and for their function in B cell maturation in GCs - when upregulated in CD4 T cells, Bcl6 initiates a program of gene expression that drives a TFH cell phenotype. Preliminary studies have demonstrated that Bcl6 is also required for downregulation of P-selectin glycoprotein ligand-1 (PSGL1), a surface protein on T cells that engages the chemokines CCL19 and CCL21 expressed in the T cell zone of secondary lymphoid organs. Indeed, TFH cells in normal and systemic autoimmune responses are uniformly PSGL1lo. These observations suggest that downregulation of PSGL1 would disengage binding to these chemokines, allowing T zone egress with migration towards the B cell follicle where TFH cells are functionally required. This would support a scenario in which Bcl6, in its role as an initiator the TFH cell developmental program, is required for both T zone egress and for development of functional TFH cells. During these studies on TFH cell development and function, the transcription factor, Ascl2, a conserved member of the Drosophila Achaete scute-like 2 complex, was also found to be selectively upregulated in this T cell subset compared to other CD4 T cell effector subsets and compared to B cells. Expression of Ascl2 also is sufficient for development of a TFH cell phenotype. Based upon these findings, two hypotheses will be addressed in this proposal. First, PSGL1 acts as a retention signal for T cells in the T zone, and that in addition to driving TFH cell development, Bcl6 expression initiates downregulation of PSGL1 as a critical first step in T cell migration to the B cell follicle and ultimately in TFH cell function. Second, the transcription factor Ascl2, like Bcl6, plays a necessary role for TFH cell development and/or function. These hypotheses will be explored in two specific aims: 1) determining the roles of Bcl6 and PSGL1 in the initial events that lead to TFH cell migration and development;2) asking if the transcription factor Ascl2 is required for TFH cell differentiation and function, and if so, a dissection of the mechanism of its T cell-specific effects in immune responses. PUBLIC HEALTH RELEVANCE: T cells that help B cells produce antibodies in the spleen and other lymphoid organs are critical for normal and autoimmune responses. In comparison to other types of T cells, less is known about the biology of these cells, including the signals that are required for their development and function. The results of the proposed studies, that utilize a combination of cellular, genetic, and imaging approaches, should help address these issues, and advance knowledge of how normal and autoimmune responses develop in humans.